Tuberculosis is the leading cause of death from infectious diseases worldwide, with increasing consequences due to the HIV epidemic. Infection and disease are caused by the intracellular bacterial pathogen Mycobacterium tuberculosis. The existing vaccine, BCG, has marginal efficacy and drug therapies are long and complex leading to noncompliance and the rapid spread of multi-drug resistant bacteria. Additionally, in the developed world, the increasing use of TNF blockade for the treatment of rheumatologic and immunological conditions has led to the activation of tuberculosis in these patients. In the face of a complex immune response, mycobacteria can persist indefinitely in granulomas, tight aggregates of highly differentiated macrophages and other immune cells. As with most infectious diseases, the exact impact of the different components of host immunity on tuberculosis infection and disease are not well understood. To better understand the host responses to tuberculous infections, we will take advantage of the fact that Mycobacterium marinum, a close relative of M. tuberculosis, is a natural pathogen of zebrafish, causing a tuberculosis-like disease with all the basic pathologic features of human tuberculosis Zebrafish are genetically tractable vertebrates that are used as models of disease and development. We have exploited the optical transparency and the genetic tractablility of zebrafish embryos and larvae to monitor and modulate host-pathogen interactions of tuberculosis in real time. Due to our ability to perform real-time imaging of the individual steps of infection, we have been able to determine the precise steps at which key host immune determinants (e.g. TNF) and bacterial virulence determinants act during pathogenesis. We will combine a variety of molecular and genetic techniques to intercept the expression of host immune functions so as to determine their impact on the individual steps and final outcomes of tuberculosis by a variety of real- time microscopy techniques.